Background

[PubMed]

Integrins are a family of cell-surface heterodimeric glycoproteins that mediate diverse biological events (e.g., cell adhesion, migration, differentiation, proliferation, and apoptosis) involving cell–cell and cell–matrix interactions (1, 2). They consist of an α and a β subunit. They are important for cell adhesion and signal transduction. On the other hand, integrins affect tumor growth, tumor invasiveness, and metastasis (3, 4). The α₂β₁ integrin receptor binds mainly collagen type I, laminins, E-cadherin, and matrix metalloproteinase 1 (5). The α₂β₁ integrin is strongly expressed on tumor cells and has been implicated in tumor progression and metastasis (6, 7). In particular, prostate cancer cells and prostate cancer stem cells express high levels of α₂β₁ integrin (8, 9). A tetrapeptide sequence consisting of Asp-Gly-Glu-Ala (DGEA) has been identified as a recognition motif used by the type I collagen to bind to α₂β₁ integrin (10). DGEA was conjugated with Cy5.5 to study in vivo biodistribution of the tracer in prostate tumor-bearing mice (11). Cy5.5-DGEA has been shown to have a high accumulation in α₂β₁-positive PC-3 human prostate tumor cells in nude mice. For positron emission tomography (PET) imaging of α₂β₁ integrin, ⁶⁴Cu-Z-E-(1,8-diamino-3,6,10,13,16,19-hexaazabicyclo(6,6,6)eicosane)-aminohexanoyl-Asp-Gly-Glu-Ala (⁶⁴Cu-Z-E(diamsar)-Ahx-DGEA) was prepared and evaluated in nude mice bearing human PC-3 prostate tumors (12).

Synthesis

[PubMed]

Z-E-(diamsar)-Ahx-DGEA peptides were obtained using solid-phase synthesis (12). The diamsar group was added to carboxy group of glutamic acid of Z-E-Ahx-DGEA peptides. The measured mass of Z-E(diamsar)-Ahx-DGEA peptides confirmed the 1:1 addition. Z-E-(diamsar)-Ahx-DGEA was incubated with [⁶⁴Cu]acetate in ammonium acetate buffer (pH 5.5) for 30 min at 23–37°C. ⁶⁴Cu-Z-E-(diamsar)-Ahx-DGEA was purified with high-performance liquid chromatography. This procedure provided a radiolabeling yield of >90%, with a specific activity of 10.7 GBq/µmol (0.29 Ci/µmol) and a radiochemical purity of >98%. ⁶⁴Cu-Z-E-(diamsar)-Ahx-DGEA was 98% intact for 48 h at 40°C in phosphate-buffered saline.

In Vitro Studies: Testing in Cells and Tissues

[PubMed]

Flow cytometry analysis showed that 99.7% of PC-3 cells, 51.4% of CWR-22 cells, and 15.6% of LNCaP cells were positive for the α₂β₁ receptors using 5-Carboxyfluorescin-DGEA (FAM-DGEA) (11). Binding of 1 µM FAM-DGEA to the three cell types was analyzed with fluorescence microscopy. PC-3 cells exhibited a higher fluorescence intensity signal than CWR-22 and LNCaP cells. The binding of FAM-DGEA to PC-3 cells was completely blocked with 20 µM DGEA. Cellular accumulation of ⁶⁴Cu-Z-E-(diamsar)-Ahx-DGEA in PC-3 cells was low, with <0.4% incubation dose at 2 h after incubation (13). No blocking studies were carried out.

Animal Studies

Human Studies

[PubMed]

No publication is currently available.

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Huang C.W., Li Z., Cai H., Chen K., Shahinian T., Conti P.S. Design, synthesis and validation of integrin alpha2beta1-targeted probe for microPET imaging of prostate cancer. Eur J Nucl Med Mol Imaging. 2011;38(7):1313–22. [PubMed: 21350963]